International Journal of Morphology
versión On-line ISSN 0717-9502
Int. J. Morphol. v.23 n.2 Temuco 2005
Int. J. Morphol.,
Foscarnet Effects on the Rat Pregnancy Outcome
Efectos del Foscarnet Sobre la Rata Preñada
*Cícero Venneri Mathias; **Manuel de Jesus Simões; **Abes Mahmed Amed; *** Ricardo Martins Oliveira-Filho; *Camila Fernandes Venneri Mathias; **Ricardo dos Santos Simões & **Luiz Kulay Júnior
* Department of Obstetrics and Ginaecology School of Medicine, Fundação ABC, São Paulo (SP), Brazil.
** Federal University of São Paulo School of Medicine (UNIFESP-EPM), São Paulo (SP), Brazil.
*** University of São Paulo Institute of Biomedical Sciences (USP), São Paulo (SP), Brazil.
SUMMARY: There are few long-term data on which to base decisions of drug management of HIV infection in pregnancy. The determination of safe medications must take into consideration the need for certain drugs and the possibility of inadvertent fetal exposure because of unplanned pregnancies. The aim of this study was to evaluate the effects of foscarnet on the entire period of rat pregnancy. Female pregnant rats were randomly assigned to four treatment groups (n = 10): one control (C) treated with the drug vehicle (bidestilled water) and three experimental groups (E1, E2 and E3) treated with 180, 360 or 720 mg/Kg of foscarnet, respectively. Rats were treated by gavage once daily. The treatment period extended from the first until the 20th day of pregnancy. Body weights were recorded weekly along this period. At term, the rats were sacrificed, the implantation sites and the number of fetuses and resorptions were recorded. The fetuses were evaluated for externally visible abnormalities under a stereomicroscope. No differences in body weights among the groups were observed; however, foscarnet-treated rats showed reduced fetal and placental weights. The incidence 137of resorptions and major malformations (shortening of limbs) in the E3 group was significantly raised. Foscarnet treatment during the entire period of rat pregnancy can produce definite toxic effects, mainly on the placental and fetal compartments.
KEY WORDS: Antiretroviral drugs; Foscarnet; Pregnancy; Rat.
RESUMEN: Foscarnet es un inhibidor de la transcriptasis reversa del HIV que actúa en la síntesis del DNA. En este trabajo evaluamos los efectos crónicos del foscarnet durante la preñez de la rata albina. Ratas preñadas fueron distribuidas aleatoriamente en cuatro grupos (n = 10 para cada grupo): uno control (C), tratadas con agua bidestilada, y tres experimentales (E1, E2 y E3), tratadas con 180, 360 o 720 mg/Kg al día de foscarnet. El fármaco y el vehículo (siempre 1 ml) fueron administrados una vez al día desde el día 0 hasta el día 20 de la gestación. Las ratas fueron pesadas semanalmente y sacrificadas al término de la preñez. No se observaron alteraciones significativas en cuanto al incremento de peso corporal entre los grupos. Sin embargo, las ratas tratadas con foscarnet (especialmente las de los grupos E2 y E3) presentaron reducciones del peso promedio de los fetos y de las respectivas placentas. La incidencia de reabsorciones y malformaciones (acortamiento de miembros) fue significativa en el grupo E3. Se concluye que la administración de foscarnet durante toda la preñez de la rata puede producir efectos tóxicos definidos, especialmente en los compartimientos placentario y fetal.
PALABRAS CLAVE: Fármacos antiretrovirales; Foscarnet; Preñez; Rata hembra.
The incidence of human immunodeficiency virus (HIV) infection is raising over the world, mainly in poor parts of Eastern Europe, Asia and sub-Saharan Africa (Bicego et al., 2003; Goodwin et al., 2004). In addition, the great number of HIV-infected pregnant women makes this disease to be an important public health concern around the world (Brettle & Leen, 1991). Therefore, special attention is given to the subject of the perinatal transmission of HIV, one of the strategies in this issue being the use of antiretroviral drug administration during pregnancy in order to prevent vertical infection (Connor et al., 1994; Tuntland et al., 1999). In fact, antiviral agents offer significant reduction in the rate of mother-to-child HIV transmission (Connor et al.; Katz, 2003). However, antiretroviral drugs occasionally induce severe side effects and may not reach their therapeutic goals in some patients, for whom the survival expectance depends upon rapid access to new antiretroviral drugs and new therapeutic strategies (Gastaut, 2003; Volberding, 2003).
Foscarnet (trisodium phosphonoformate) is a synthetic pyrophosphate analogue which effectively acts against citomegalovirus and herpesvirus, being also an alternative against the HIV reverse transcriptase. Its antiviral activity is exerted through a selective inhibition at the pyrophosphate binding site of virus-specific DNA polymerases, at concentrations that do not affect host cellular DNA polymerases. Unfortunately, there are no adequate or well- controlled studies in pregnant women using foscarnet. It is known that its administration up to 75 mg/day can cause some rat fetal malformations, such as skeletal anomalies (Watts, 1992).
Since the chronic effects of foscarnet during pregnancy were not fully studied as yet, this paper aims to contribute to this knowledge by evaluating the effects of its administration to pregnant rats in three dosage levels during the entire period of gestation.
MATERIAL AND METHOD
The experimental protocols were approved by the local Committee of Ethics on Animal Experimentation. Adult virgin female EPM-1 Wistar rats, weighing about 200 g, were selected after three regular consecutive estrous cycles and kept under specific pathogen-free conditions. The animals were allowed free access to a standard Purina® pelleted diet and water and were maintained at 1922C, in a 12-h light/dark cycle (lights on at 7:00 a. m.). The rats were mated in the proportion of 2 females for one male, during 2 h. The day 'zero' of pregnancy was considered to be the first 24-h period after the mating by the finding of spermatozoa in vaginal smears (Hamilton & Wolfe, 1938).
Forty pregnant rats were then randomly divided into four groups with ten rats each, one control (C) and three experimental drug-treatment groups (E1, E2 and E3). Control animals were given the drug vehicle (bidestilled water) from day 0 up to the 20th day of pregnancy. The experimental groups were given 180 mg/Kg (E1), 360 mg/Kg (E2) or 720 mg/Kg (E3) per day of foscarnet, respectively, during the entire period of pregnancy. The drug (foscarnet trisodium salt, hexahydrated, Astra Laboratory) was purchased as a solution for intravenous administration containing 24 mg/ml of foscarnet. Drug and vehicle administrations were done once a day (at 10:00h a.m.) by intraperitoneal route. Body weights were recorded on the «zero» and the 7th, 14th and 20th days of pregnancy.
At term (20th day), the rats were killed by decapitation under deep anesthesia using 1 mg/Kg ketamin. Upon laparotomy and opening of the uterine horns, the following items were recorded: number of implantations and resorptions, number of placentae, placental and fetal weights, and number of intrauterine deaths. Fetal malformations were studied with a stereomicroscope.
A resorption is herein defined as the finding of endometrial sites with an appended amorphous mass without a fetus. The number of implantations is defined as the results of the total number of placentae with fetuses plus the total number of resorptions.
The results were analyzed by one-way analysis of variance (ANOVA) and the Tukey-Kramer's multiple comparisons test. A 2.01 version of GraphPad InStatTM software was used for this purpose. The differences were considered statistically significant when p<0.05. The Fisher's exact test was used for analysis of contingency tables.
RESULTS AND DISCUSSION
The weight of all females at mating averaged 213.57 ± 12.05 g (mean ± SD). Table I summarizes the rat body weight gain during the experiment. At term, the mean body weight of all animals averaged 291.67 ± 20.50 g, and no significant differences were noticed among the groups (Fig. 1). The results of pregnancy outcome after treatments are seen in Table II. The mean fetal weight in the E3 group was significantly decreased regarding the other groups (p<0.001). The placental weight was reduced in the group treated with the largest dose of foscarnet (group E3), and this coincided with a significantly increased number of foetal resorptions (Table II). In addition, there has been a significant number of foetal limb shortenings in that group.
One aspect of concern during antiviral therapy with foscarnet is that, unfortunately, there are no adequate or well-controlled studies in pregnant women using foscarnet; moreover, there are reports of nephrotoxicity (36%) and liver damage (5%) in humans (Sjovall et al., 1988, 1989; Fanning et al., 1990; Jayaweera, 1997). The administration of this drug up to 75 mg/day per day may cause some rat fetal malformations, such as skeletal anomalies (Watts).
Since no reports in the literature were found about the effects foscarnet on the body weight gain of pregnant rats, this aspect was examined in our animals. Table I and Fig. 1 show the body weight gain of rats during pregnancy throughout the experimental period. Similarly to what occurs in normal rats, it should be noticed that all groups reached their fastest weight gain in the last third of pregnancy, but no differences were detected with regard to the control group.
The drug administration protocol used herein corresponded to a foetal exposition to foscarnet during the entire gestational period, comprising the phases of cell hyperplasia, hyperplasia/hypertrophy and hypertrophy (Winick, 1970).
In the course of foscarnet use for the treatment of cytomegalovirus retinitis in patients with AIDS, Jacobson et al. (1989) and Lehoang et al. (1989) reported the occurrence of hypomagnesemia, hypocalemia, hypocalcemia and hypophosphatemia. Accordingly, Paspaliaris et al. (1993) showed that foscarnet inhibits vascular smooth muscle contraction by inhibiting calcium release from intracellular stores, and it seems that the foscarnet molecule binds preferentially magnesium ions (Noormohamed et al., 1996). The actions exerted by foscarnet at the mineral metabolism level could help to explain, at least in part, why pregnant rats treated during the entire period of gestation display deleterious effects, mainly at placental and foetal compartments. Accordingly, it is highly conceivable that the shortening of foetal limbs seen in the group E3 may relate to the interference of foscarnet with calcium ions.
Fig. 1. Evolution of body weight gain during pregnancy in rats, control (C) or treated during the entire period of pregnancy with foscarnet (E1 = 180 mg/Kg; E2 = 360 mg/Kg; E3 = 720 mg/Kg once a day, by gavage). Values are mean ± SEM (n = 10 for every group) of percentual increments with regard to the initial weight (pregnancy day zero). No significant differences were detected among the groups.
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Prof. Dr. Cícero Venneri Mathias
Departamento de Obstetrícia.
Universidade Federal de São Paulo UNIFRESP/EPM
Rua Dr. Esdras Pacheco Ferreira, 143 Vila Nova Conceição
São Paulo, SP. BRAZIL
Received : 11-02-2005